High-Pass Filtering of Input Signals by the Ih Current in a Non-Spiking Neuron, the Retinal Rod Bipolar Cell

Hyperpolarization–activated cyclic nucleotide–sensitive (HCN) channels mediate the If current in heart and Ih throughout the nervous system. In spiking neurons Ih participates primarily in different forms of rhythmic activity. Little is known, however, about its role in neurons operating with graded potentials as in the retina, where all four channel isoforms are expressed. Intriguing evidence for an involvement of Ih in early visual processing are the side effects reported, in dim light or darkness, by cardiac patients treated with HCN inhibitors. Moreover, electroretinographic recordings indicate that these drugs affect temporal processing in the outer retina. Here we analyzed the functional role of HCN channels in rod bipolar cells (RBCs) of the mouse. Perforated–patch recordings in the dark–adapted slice found that RBCs exhibit Ih, and that this is sensitive to the specific blocker ZD7288. RBC input impedance, explored by sinusoidal frequency–modulated current stimuli (0.1–30 Hz), displays band–pass behavior in the range of Ih activation. Theoretical modeling and pharmacological blockade demonstrate that high–pass filtering of input signals by Ih, in combination with low–pass filtering by passive properties, fully accounts for this frequency–tuning. Correcting for the depolarization introduced by shunting through the pipette–membrane seal, leads to predict that in darkness Ih is tonically active in RBCs and quickens their responses to dim light stimuli. Immunohistochemistry targeting candidate subunit isoforms HCN1–2, in combination with markers of RBCs (PKC) and rod–RBC synaptic contacts (bassoon, mGluR6, Kv1.3), suggests that RBCs express HCN2 on the tip of their dendrites. The functional properties conferred by Ih onto RBCs may contribute to shape the retina's light response and explain the visual side effects of HCN inhibitors

Bordetella Adenylate Cyclase Toxin Mobilizes Its β2 Integrin Receptor into Lipid Rafts to Accomplish Translocation across Target Cell Membrane in Two Steps

Bordetella adenylate cyclase toxin (CyaA) binds the αMβ2 integrin (CD11b/CD18, Mac-1, or CR3) of myeloid phagocytes and delivers into their cytosol an adenylate cyclase (AC) enzyme that converts ATP into the key signaling molecule cAMP. We show that penetration of the AC domain across cell membrane proceeds in two steps. It starts by membrane insertion of a toxin ‘translocation intermediate’, which can be ‘locked’ in the membrane by the 3D1 antibody blocking AC domain translocation. Insertion of the ‘intermediate’ permeabilizes cells for influx of extracellular calcium ions and thus activates calpain-mediated cleavage of the talin tether. Recruitment of the integrin-CyaA complex into lipid rafts follows and the cholesterol-rich lipid environment promotes translocation of the AC domain across cell membrane. AC translocation into cells was inhibited upon raft disruption by cholesterol depletion, or when CyaA mobilization into rafts was blocked by inhibition of talin processing. Furthermore, CyaA mutants unable to mobilize calcium into cells failed to relocate into lipid rafts, and failed to translocate the AC domain across cell membrane, unless rescued by Ca2+ influx promoted in trans by ionomycin or another CyaA protein. Hence, by mobilizing calcium ions into phagocytes, the ‘translocation intermediate’ promotes toxin piggybacking on integrin into lipid rafts and enables AC enzyme delivery into host cytosol

A Resource for Discovering Specific and Universal Biomarkers for Distributed Stem Cells

Specific and universal biomarkers for distributed stem cells (DSCs) have been elusive. A major barrier to discovery of such ideal DSC biomarkers is difficulty in obtaining DSCs in sufficient quantity and purity. To solve this problem, we used cell lines genetically engineered for conditional asymmetric self-renewal, the defining DSC property. In gene microarray analyses, we identified 85 genes whose expression is tightly asymmetric self-renewal associated (ASRA). The ASRA gene signature prescribed DSCs to undergo asymmetric self-renewal to a greater extent than committed progenitor cells, embryonic stem cells, or induced pluripotent stem cells. This delineation has several significant implications. These include: 1) providing experimental evidence that DSCs in vivo undergo asymmetric self-renewal as individual cells; 2) providing an explanation why earlier attempts to define a common gene expression signature for DSCs were unsuccessful; and 3) predicting that some ASRA proteins may be ideal biomarkers for DSCs. Indeed, two ASRA proteins, CXCR6 and BTG2, and two other related self-renewal pattern associated (SRPA) proteins identified in this gene resource, LGR5 and H2A.Z, display unique asymmetric patterns of expression that have a high potential for universal and specific DSC identification

The Herpes Simplex Virus-1 Transactivator Infected Cell Protein-4 Drives VEGF-A Dependent Neovascularization

Herpes simplex virus-1 (HSV-1) causes lifelong infection affecting between 50 and 90% of the global population. In addition to causing dermal lesions, HSV-1 is a leading cause of blindness resulting from recurrent corneal infection. Corneal disease is characterized by loss of corneal immunologic privilege and extensive neovascularization driven by vascular endothelial growth factor-A (VEGF-A). In the current study, we identify HSV-1 infected cells as the dominant source of VEGF-A during acute infection, and VEGF-A transcription did not require TLR signaling or MAP kinase activation. Rather than being an innate response to the pathogen, VEGF-A transcription was directly activated by the HSV-1 encoded immediate early transcription factor, ICP4. ICP4 bound the proximal human VEGF-A promoter and was sufficient to promote transcription. Transcriptional activation also required cis GC-box elements common to the VEGF-A promoter and HSV-1 early genes. Our results suggest that the neovascularization characteristic of ocular HSV-1 disease is a direct result of HSV-1's major transcriptional regulator, ICP4, and similarities between the VEGF-A promoter and those of HSV-1 early genes

Synergistic Interactions between the NS3hel and E Proteins Contribute to the Virulence of Dengue Virus Type 1

Dengue virus constitutes a significant public health problem in tropical regions of the world. Despite the high morbidity and mortality of this infection, no effective antiviral drugs or vaccines are available for the treatment or prevention of dengue infections. The profile of clinical signs associated with dengue infection has changed in recent years with an increase in the number of episodes displaying unusual signs. We use reverse genetics technology to engineer DENV-1 viruses with subsets of mutations previously identified in highly neurovirulent strains to provide insights into the molecular mechanisms underlying dengue neuropathogenesis. We found that single mutations affecting the E and NS3hel proteins, introduced in a different genetic context, had a synergistic effect increasing DENV replication capacity in human and mosquito derived cells in vitro. We also demonstrated correlations between the presence of these mutations and viral replication efficiency, viral loads, the induction of innate immune response genes and pathogenesis in a mouse model. These results should improve our understanding of the DENV-host cell interaction and contribute to the development of effective antiviral strategies

Diversity, Phylogeny and Expression Patterns of Pou and Six Homeodomain Transcription Factors in Hydrozoan Jellyfish Craspedacusta sowerbyi

Formation of all metazoan bodies is controlled by a group of selector genes including homeobox genes, highly conserved across the entire animal kingdom. The homeobox genes from Pou and Six classes are key members of the regulation cascades determining development of sensory organs, nervous system, gonads and muscles. Besides using common bilaterian models, more attention has recently been targeted at the identification and characterization of these genes within the basal metazoan phyla. Cnidaria as a diploblastic sister group to bilateria with simple and yet specialized organs are suitable models for studies on the sensory organ origin and the associated role of homeobox genes. In this work, Pou and Six homeobox genes, together with a broad range of other sensory-specific transcription factors, were identified in the transcriptome of hydrozoan jellyfish Craspedacusta sowerbyi. Phylogenetic analyses of Pou and Six proteins revealed cnidarian-specific sequence motifs and contributed to the classification of individual factors. The majority of the Craspedacusta sowerbyi Pou and Six homeobox genes are predominantly expressed in statocysts, manubrium and nerve ring, the tissues with sensory and nervous activities. The described diversity and expression patterns of Pou and Six factors in hydrozoan jellyfish highlight their evolutionarily conserved functions. This study extends the knowledge of the cnidarian genome complexity and shows that the transcriptome of hydrozoan jellyfish is generally rich in homeodomain transcription factors employed in the regulation of sensory and nervous functions

Myocardial tagging by Cardiovascular Magnetic Resonance: evolution of techniques--pulse sequences, analysis algorithms, and applications

Cardiovascular magnetic resonance (CMR) tagging has been established as an essential technique for measuring regional myocardial function. It allows quantification of local intramyocardial motion measures, e.g. strain and strain rate. The invention of CMR tagging came in the late eighties, where the technique allowed for the first time for visualizing transmural myocardial movement without having to implant physical markers. This new idea opened the door for a series of developments and improvements that continue up to the present time. Different tagging techniques are currently available that are more extensive, improved, and sophisticated than they were twenty years ago. Each of these techniques has different versions for improved resolution, signal-to-noise ratio (SNR), scan time, anatomical coverage, three-dimensional capability, and image quality. The tagging techniques covered in this article can be broadly divided into two main categories: 1) Basic techniques, which include magnetization saturation, spatial modulation of magnetization (SPAMM), delay alternating with nutations for tailored excitation (DANTE), and complementary SPAMM (CSPAMM); and 2) Advanced techniques, which include harmonic phase (HARP), displacement encoding with stimulated echoes (DENSE), and strain encoding (SENC). Although most of these techniques were developed by separate groups and evolved from different backgrounds, they are in fact closely related to each other, and they can be interpreted from more than one perspective. Some of these techniques even followed parallel paths of developments, as illustrated in the article. As each technique has its own advantages, some efforts have been made to combine different techniques together for improved image quality or composite information acquisition. In this review, different developments in pulse sequences and related image processing techniques are described along with the necessities that led to their invention, which makes this article easy to read and the covered techniques easy to follow. Major studies that applied CMR tagging for studying myocardial mechanics are also summarized. Finally, the current article includes a plethora of ideas and techniques with over 300 references that motivate the reader to think about the future of CMR tagging

A chromosome conformation capture ordered sequence of the barley genome

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